Project Summary/Abstract A complex and diverse community of bacteria reside on human skin. Abundant experimental evidence now exists in cell culture systems and mouse models that shows many of these commensal bacteria residing on healthy subjects can be beneficial. These bacteria perform essential functions such as inhibiting survival of pathogenic bacteria, limiting skin inflammation and enhancing skin innate immune defense. Therefore, it has been hypothesized that the bacteria that normally inhabit human skin could be exploited to benefit us. However, clinical proof in humans of this hypothesis is lacking because of the lack of a rational controlled approach to test this. This time-sensitive mechanistic ancillary study to our ongoing exploratory and developmental clinical trial seeks to study the mechanist effect of potential benefits of transplant of commensal bacteria part of the microbiome on human skin (autologous microbial transplant ? AMT). Based on exciting progress of the efficacy of the AMT to reduce colonization of atopic dermatitis lesional skin by Staphylococcus aureus we propose to utilize a novel ultra-low input protocol to gain insight into the functionality of the AMT by analyzing the transcriptional response of the resident skin microbiome, S. aureus, and the AMT- bacteria over time. As demonstrated before, we will use scientific rigor to generate accurate and reproducible results and share all data as laid out in our data sharing plan. If successful, the study will provide essential data to design future experiments to measure the therapeutic potential of a customized microbial approach to enhance cutaneous host defense. Insight into the function and mechanism of AMT treatment on S. aureus and the skin microbiome are crucial for improving and optimizing such a therapy so future studies can be designed to focus on the overall therapeutic value of the skin microbiome. This proposal will provide answers to key unknown questions about the microbiome of human skin that will be relevant to a wide range of skin disorders. Our specific aims are: Aim 1: Determine the transcriptional response of the resident AD skin microbiome after autologous microbiome transplantation AMT. Aim 2: Evaluate the transcriptional profile of S. aureus before, during and after AMT. Aim 3: Measure the transcriptional response of the transplanted bacteria after colonization of AD skin.